Our-long term objective has been to study the properties of C3, C4, and C5 that are related to their structural integrity and functionality. These proteins are components of the complement system, whose apparent main function is to help defend the human body against bacteria, viruses, yeast and foreign particles or substances. All three proteins have to be activated and fragmented to fulfill a variety of their physiological functions. Most of the fragments have the capacity to trigger cellular responses, some may participate even in an immune regulation. An additional stability to functional forms of C3, C4, and C5, or their fragments, is being conferred by disulfide bonds. The specific aims of this project are: (1) To demonstrate the effect of distribution of disulfide linkages on conformation of the domains of C3 and C4, and to localize interchain, inter- and intradomains linkages in C3, C4, and C5. Determination of disulfide-bonding patterns will allow to understand the spacial arrangements within the polypeptide chains, and should provide another evidence for the importance of disulfide bonds for the structural and functional integrity in homologous proteins. The study of the interaction of highly cross- linked domains, and other domains, with physiological ligands will be attempted. (2) To elucidate the fragmentation pathway of the fluid-phase C3b, the activated form of C3. The physiologically relevant cleavage products generated in the absence of acceptor surfaces will be isolated by immunoaffinity chromatography and characterized by SDS-PAGE and by N-terminal sequence analysis. We propose to identify the enzyme responsible for the cleavage of the fluid-phase iC3b into C3c and C3d-containing fragments. Biological activities of the fluid-phase fragments, specifically those of C3d,g dimer and C3c, will be investigated.